Catalytic Fields as a Tool to Analyze Enzyme Reaction Mechanism Variants and Reaction Steps.

Catalytic fields representing the topology of the optimal molecular environment charge distribution that reduces the activation barrier have been used to examine alternative reaction variants and to determine the role of conserved catalytic residues for two consecutive reactions catalyzed by the same enzyme. Until now, most experimental and conventional top-down theoretical studies employing QM/MM or ONIOM methods have focused on the role of enzyme electric fields acting on broken bonds of reactants. In contrast, our bottom-up approach dealing with a small reactant and transition-state model allows the analysis of the opposite effects: how the catalytic field resulting from the charge redistribution during the enzyme reaction acts on conserved amino acid residues and contributes to the reduction of the activation barrier. This approach has been applied to the family of histidyl tRNA synthetases involved in the translation of the genetic code into the protein amino acid sequence. Activation energy changes related to conserved charged amino acid residues for 12 histidyl tRNA synthetases from different biological species allowed to compare on equal footing the catalytic residues involved in ATP aminoacylation and tRNA charging reactions and to analyze different reaction mechanisms proposed in the literature. A scan of the library of atomic multipoles for amino acid side-chain rotamers within the catalytic field pointed out the change in the Glu83 conformation as the critical catalytic effect, providing, at low computational cost, insight into the electrostatic preorganization of the enzyme catalytic site at a level of detail that has not yet been accessible in conventional experimental or theoretical methods. This opens the way for rational reverse biocatalyst design at a very limited computational cost without resorting to empirical methods.

Extreme Catalytic Power of Ketosteroid Isomerase Related to the Reversal of Proton Dislocations in Hydrogen-Bond Network.

Dynamic electrostatic catalytic field (DECF) vectors derived from transition state and reactant wavefunctions for the two-step reaction occurring within ketosteroid isomerase (KSI) have been calculated using MP2/aug-cc-pVTZ and lower theory levels to determine the magnitude of the catalytic effect and the optimal directions of proton transfers in the KSI hydrogen-bond network. The most surprising and meaningful finding is that the KSI catalytic activity is enhanced by proton dislocations proceeding in opposite directions for each of the two consecutive reaction steps in the same hydrogen network. Such a mechanism allows an ultrafast switching of the catalytic proton wire environment, possibly related to the exceptionally high KSI catalytic power.

Bottom-Up Nonempirical Approach To Reducing Search Space in Enzyme Design Guided by Catalytic Fields.

Currently developed protocols of theozyme design still lead to biocatalysts with much lower catalytic activity than enzymes existing in nature, and, so far, the only avenue of improvement was the in vitro laboratory-directed evolution (LDE) experiments. In this paper, we propose a different strategy based on "reversed" methodology of mutation prediction. Instead of common "top-down" approach, requiring numerous assumptions and vast computational effort, we argue for a "bottom-up" approach that is based on the catalytic fields derived directly from transition state and reactant complex wave functions. This enables direct one-step determination of the general quantitative angular characteristics of optimal catalytic site and simultaneously encompasses both the transition-state stabilization (TSS) and ground-state destabilization (GSD) effects. We further extend the static catalytic field approach by introducing a library of atomic multipoles for amino acid side-chain rotamers, which, together with the catalytic field, allow one to determine the optimal side-chain orientations of charged amino acids constituting the elusive structure of a preorganized catalytic environment. Obtained qualitative agreement with experimental LDE data for Kemp eliminase KE07 mutants validates the proposed procedure, yielding, in addition, a detailed insight into possible dynamic and epistatic effects.

Predicting substituent effects on activation energy changes by static catalytic fields.

Catalytic fields illustrate topology of the optimal charge distribution of a molecular environment reducing the activation energy for any process involving barrier crossing, like chemical reaction, bond rotation etc. Until now, this technique has been successfully applied to predict catalytic effects resulting from intermolecular interactions with individual water molecules constituting the first hydration shell, aminoacid mutations in enzymes or Si→Al substitutions in zeolites. In this contribution, hydrogen to fluorine (H→F) substitution effects for two model reactions have been examined indicating qualitative applicability of the catalytic field concept in the case of systems involving intramolecular interactions. Graphical abstract Hydrogen to fluorine (H→F) substitution effects on activation energy in [kcal/mol].

Application of a simple quantum chemical approach to ligand fragment scoring for Trypanosoma brucei pteridine reductase 1 inhibition.

There is a need for improved and generally applicable scoring functions for fragment-based approaches to ligand design. Here, we evaluate the performance of a computationally efficient model for inhibitory activity estimation, which is composed only of multipole electrostatic energy and dispersion energy terms that approximate long-range ab initio quantum mechanical interaction energies. We find that computed energies correlate well with inhibitory activity for a compound series with varying substituents targeting two subpockets of the binding site of Trypanosoma brucei pteridine reductase 1. For one subpocket, we find that the model is more predictive for inhibitory activity than the ab initio interaction energy calculated at the MP2 level. Furthermore, the model is found to outperform a commonly used empirical scoring method. Finally, we show that the results for the two subpockets can be combined, which suggests that this simple nonempirical scoring function could be applied in fragment-based drug design.

Are Various σ-Hole Bonds Steered by the Same Mechanisms?

Representative Lewis acid-Lewis base complexes linked by tetrel, pnicogen, chalcogen, and halogen bonds have been studied within the quantum theory of atoms in molecules (QTAIM) approach and the hybrid variation-perturbation theory (HVPT) to analyze possible relationships between these σ-hole dimers. Results obtained at the MP2/aug-cc-pVTZ level indicate numerous correlations similar to hydrogen-bonded systems.

Rapid Estimation of Catalytic Efficiency by Cumulative Atomic Multipole Moments: Application to Ketosteroid Isomerase Mutants.

We propose a simple atomic multipole electrostatic model to rapidly evaluate the effects of mutation on enzyme activity and test its performance on wild-type and mutant ketosteroid isomerase. The predictions of our atomic multipole model are similar to those obtained with symmetry-adapted perturbation theory at a fraction of the computational cost. We further show that this approach is relatively insensitive to the precise amino acid side chain conformation in mutants and may thus be useful in computational enzyme (re)design.

Universal short-range ab initio atom-atom potentials for interaction energy contributions with an optimal repulsion functional form.

The repulsion term in conventional force fields constitutes a major source of error. Assuming that this could originate from a too simple analytical functional form, we analyzed various analytical functions using ab initio exchange component values as a reference and obtained (α + ß R (-1))exp(-γ R) as the optimal form to represent the repulsion term. Universal exchange, delocalization, and electrostatic penetration potentials approximating the corresponding interaction energy components defined within hybrid variation-perturbation theory (HVPT) were derived using as a reference a training set of 660 biomolecular complexes. The electrostatic multipole term was calculated using cumulative atomic multipole moments, whereas correlation contribution including dispersion term and first-order correlation correction was estimated from nonempirical D a s functions derived by Pernal et al. The resulting non-empirical atom-atom potentials (NEAAP) were tested for several urokinase-inhibitor complexes yielding improved docking results.

Physical Nature of Fatty Acid Amide Hydrolase Interactions with Its Inhibitors: Testing a Simple Nonempirical Scoring Model.

Fatty acid amide hydrolase (FAAH) is an enzyme responsible for the deactivating hydrolysis of fatty acid ethanolamide neuromodulators. FAAH inhibitors have gained considerable interest due to their possible application in the treatment of anxiety, inflammation, and pain. In the context of inhibitor design, the availability of reliable computational tools for predicting binding affinity is still a challenging task, and it is now well understood that empirical scoring functions have several limitations that in principle could be overcome by quantum mechanics. Herein, systematic ab initio analyses of FAAH interactions with a series of inhibitors belonging to the class of the N-alkylcarbamic acid aryl esters have been performed. In contrast to our earlier studies of other classes of enzyme-inhibitor complexes, reasonable correlation with experimental results required us to consider correlation effects along with electrostatic term. Therefore, the simplest comprehensive nonempirical model allowing for qualitative predictions of binding affinities for FAAH ligands consists of electrostatic multipole and second-order dispersion terms. Such a model has been validated against the relative stabilities of the benchmark S66 set of biomolecular complexes. As it does not involve parameters fitted to experimentally derived data, this model offers a unique opportunity for generally applicable inhibitor design and virtual screening.

Alkaline hydrolysis of organophosphorus pesticides: the dependence of the reaction mechanism on the incoming group conformation.

The fundamental mechanism of organophosphate hydrolysis is the subject of a growing interest resulting from the need for safe disposal of phosphoroorganic pesticides. Herein, we present a detailed ab initio study of the gas-phase mechanisms of alkaline hydrolysis of P-O and P-S bonds in a number of organophosphorus pesticides, including paraoxon, methyl parathion, fenitrothion, demeton-S, acephate, phosalone, azinophos-ethyl, and malathion. Our main finding is that the incoming group conformation influences the mechanism of decomposition of organophosphate and organothiophosphate compounds. Depending on the orientation of the attacking nucleophile, hydrolysis reaction might follow either a multistep pathway characterized by the presence of a pentavalent intermediate or a one-step mechanism proceeding through a single transition state. Despite a widely accepted view of the phosphotriester P-O bonds being decomposed exclusively via a direct-displacement mechanism, the occurrence of alternative, qualitatively distinct reaction pathways was confirmed for alkaline hydrolysis of both P-O and P-S bonds. As the pesticides included in our quantum chemical analysis involve organophosphate, phosphorothioate, and phosphorodithioate compounds, the influence of oxygen to sulfur substitution on the structural and energetic characteristics of the hydrolysis pathway is also discussed.

Physical nature of intermolecular interactions in [BMIM][PF6] ionic liquid.

The intermolecular interaction energy in a popular ionic liquid, [BMIM][PF6] is analyzed using the Hybrid Variation-Perturbation Theory approach. The analysis is performed on a sample of configurations from molecular dynamics simulation, instead of minimized structures. The interaction energy components are quantified, showing that the electrostatics is the dominating but not the only important term. It is found that two- and three-body electron delocalization components also contribute to the stabilization of the complexes; however, these interactions vanish beyond the first coordination sphere. The presented study shows a systematic way to obtain the amount of physically meaningful components of the interaction energy, which possibly could be related to macroscopic properties of ionic liquids (e.g., viscosity, melting point) or electron transfer in ionic liquids.

Low cost prediction of relative stabilities of hydrogen bonded complexes from atomic multipole moments for overly short intermolecular distances.

The relative stability of biologically relevant, hydrogen bonded complexes with shortened distances can be assessed at low cost by the electrostatic multipole term alone more successfully than by ab initio methods. These results imply that atomic multipole moments may help improve ligand-receptor ranking predictions, particularly in cases where accurate structural data are not available.

Nonempirical energetic analysis of reactivity and covalent inhibition of fatty acid amide hydrolase.

Fatty acid amide hydrolase (FAAH) is a member of the amidase signature family and is responsible for the hydrolytic deactivation of fatty acid amide neuromodulators, such as anandamide. FAAH carries an unusual catalytic triad consisting of Lys-Ser-Ser, which uniquely enables the enzyme to cleave amides and esters at similar rates. The acylation of 9Z-octadecenamide (oleamide, a FAAH reference substrate) has been widely investigated by computational methods, and those have shown that conformational fluctuations of the active site affect the reaction barrier. Empirical descriptors have been devised to provide a possible mechanistic explanation for such conformational effects, but a first-principles understanding is still missing. A comparison of FAAH acylation with a reference reaction in water suggests that transition-state stabilization is crucial for catalysis because the activation energy barrier falls by 6 kcal/mol in the presence of the active site. With this in mind, we have analyzed the enzymatic reaction using the differential transition-state stabilization (DTSS) approach to determine key active-site residues for lowering the barrier. We examined several QM/MM structures at the MP2 level of theory and analyzed catalytic effects with a variation-perturbation partitioning of the interaction energy into electrostatic multipole and penetration, exchange, delocalization, and correlation terms. Three residues - Thr236, Ser218, and one water molecule - appear to be essential for the stabilization of the transition state, a conclusion that is also reflected by catalytic fields and agrees with site-directed mutagenesis data. An analogous analysis for URB524, URB618, and URB694 (three potent representatives of covalent, carbamate-based FAAH inhibitors) confirms the importance of the residues involved in oleamide acylation, providing insight for future inhibitor design.

The Ethidium-UA/AU Intercalation Site: Effect of Model Fragmentation and Backbone Charge State.

We report a systematic analysis of the intermolecular interactions of cationic ethidium intercalated into a UA/AU step of RNA for a single conformation based on crystallographic coordinates. Interaction energies at the MP2/6-31G** level were partitioned into electrostatic, exchange, delocalization, and correlation components. Various pairwise interaction models built from chemically intuitive fragments reproduce within a few percent values obtained when treating the intercalation site as a whole. Gas phase results are very sensitive to the charge state of the two phosphate groups, with the electrostatic term nearly tripling when the counterions are removed. But this is largely compensated by solvation, an effect represented here within the polarizable continuum model. In a few cases, more diffuse and larger basis sets as well as QCISD(T) corrections were applied in an effort to estimate plausible ethidium-nucleobase electron correlation effects.

Physical nature of intermolecular interactions within cAMP-dependent protein kinase active site: differential transition state stabilization in phosphoryl transfer reaction.

The origin of enzyme catalytic activity may be effectively explored within the nonempirical theory of intermolecular interactions. The knowledge of electrostatic, exchange, delocalization, and correlation components of the transition state and substrates stabilization energy arising from each enzyme active site residue allows to examine the most essential physical effects involved in enzymatic catalysis. Consequently, one can build approximate models of the catalytic activity in a systematic and legitimate manner. Whenever the dominant role of electrostatic interactions is recognized or assumed, the properties of an optimal catalytic environment could be simply generalized and visualized by means of catalytic fields that, in turn, aids the design of new catalysts. Differential transition state stabilization (DTSS) methodology has been applied herein to the phosphoryl transfer reaction catalyzed by cAMP-dependent protein kinase (PKA). The MP2 results correlate well with the available experimental data and theoretical findings indicating that Lys72, Asp166, and the two magnesium ions contribute -22.7, -13.3, -32.4, and -15.2 kcal/mol to differential transition state stabilization, respectively. Although all interaction energy components except that of electron correlation contribution are meaningful, the first-order electrostatic term correlates perfectly with MP2 catalytic activity. Catalytic field technique was also employed to visualize crucial electrostatic features of an ideal catalyst and to compare the latter with the environment provided by PKA active site. The map of regional electronic chemical potential was used to analyze the unfavorable catalytic effect of Lys168. It was found that locally induced polarization of TS atoms thermodynamically destabilizes electrons, pulling them to regions displaying higher electronic chemical potential.

Gas-phase mechanisms of degradation of hazardous organophosphorus compounds: do they follow a common pattern of alkaline hydrolysis reaction as in phosphotriesterase?

A comprehensive ab initio analysis of the gas-phase mechanisms of alkaline hydrolysis for a number of phosphotriesterase substrates--O,O-diisopropyl phosphorofluoridate (DFP), O-isopropyl methyl phosphonofluoridate, O,O-diethyl p-nitrophenyl phosphate (paraoxon), O,O-diethyl p-nitrophenyl thiophosphate (parathion), N-acetyl phosphoramidothioate (acephate), O,O-diethyl S-2-ethylthioethyl phosphorothioate (demeton-S) and O-ethyl N,N-dimethyl phosphoramidocyanidate--has been presented herein. The results indicate that, although an associative mechanism of alkaline hydrolysis is followed by all these compounds, P-F and P-CN bonds are cleaved according to the multistep addition-elimination scheme, whereas the breakage of P-O and P-S bonds appears to be consistent with the one-step direct-displacement mechanism. Of the two alternative reaction pathways present in all those cases (except of acephate), the most probable one involves the proton from a nucleophilic hydroxide experiencing an additional stabilization by the phosphoryl oxygen atom.

Ab initio multireference study of Hetero-Diels-Alder reaction of buta-1,3-diene with alkyl glyoxylates.

Hetero-Diels-Alder (HDA) reaction of methyl glyoxylate with buta-1,3-diene has been investigated using multireference methods (complete active space SCF and multi-reference perturbation theory) and compared with several single-reference methods (including DFT) often used in calculations of catalysed [4+2] cycloadditions. Concerted and stepwise mechanisms, found in the literature, are compared. It is shown, that the stepwise mechanism may be a result of choosing unbalanced active space. Such choice leads to very close singlet and triplet states in the intermediate geometry - an artificial effect, that disappears if properly balanced active space is used (here, we use active space of 12 orbitals and 12 electrons). Conclusions concerning the mechanism and usefulness of the applied methodology are drawn, which might be important for theoretical investigation of stereoselectivity and specificity of catalysts for the HDA reaction.

Intriguing relations of interaction energy components in stacked nucleic acids.

Major components of the interaction energy that define several approximate levels starting from second order Möller-Plesset theory were studied for 58 stacked nucleic acid dimers. They included typical B-DNA and A-DNA structures, and selected published geometries. A survey of the various terms yields an unexpected correlation between the Pauli exchange and dispersion or correlation terms, which holds for each class of similar planar geometries and for various basis sets. The geometries that exhibit these correlations span a specific range of molecular overlaps when compared to a model benzene-pyridine stacked dimer. Also, the relationship between electrostatic interactions and MP2 stabilization energies reported earlier is confirmed and a prediction interval of practical relevance is estimated.